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基本情報
登録情報 | データベース: PDB / ID: 6ebk | ||||||
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タイトル | The voltage-activated Kv1.2-2.1 paddle chimera channel in lipid nanodiscs | ||||||
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![]() | MEMBRANE PROTEIN / transport protein / potassium channel / lipid nanodisc | ||||||
機能・相同性 | ![]() optic nerve structural organization / pinceau fiber / regulation of action potential / Voltage gated Potassium channels / voltage-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / potassium channel complex / NADPH oxidation / potassium ion export across plasma membrane / paranodal junction / regulation of circadian sleep/wake cycle, non-REM sleep ...optic nerve structural organization / pinceau fiber / regulation of action potential / Voltage gated Potassium channels / voltage-gated monoatomic ion channel activity involved in regulation of postsynaptic membrane potential / potassium channel complex / NADPH oxidation / potassium ion export across plasma membrane / paranodal junction / regulation of circadian sleep/wake cycle, non-REM sleep / axon initial segment / regulation of protein localization to cell surface / corpus callosum development / aldo-keto reductase (NADPH) activity / voltage-gated monoatomic ion channel activity involved in regulation of presynaptic membrane potential / juxtaparanode region of axon / regulation of potassium ion transmembrane transport / delayed rectifier potassium channel activity / outward rectifier potassium channel activity / 酸化還元酵素; CH-OHの結合に対し酸化酵素として働く; NAD又はNADPを用いる / myoblast differentiation / neuromuscular process / Neutrophil degranulation / optic nerve development / neuronal cell body membrane / regulation of dopamine secretion / voltage-gated potassium channel activity / kinesin binding / calyx of Held / lamellipodium membrane / potassium channel regulator activity / neuronal action potential / hematopoietic progenitor cell differentiation / axon terminus / potassium ion transmembrane transport / voltage-gated potassium channel complex / extrinsic component of cytoplasmic side of plasma membrane / sensory perception of pain / protein localization to plasma membrane / postsynaptic density membrane / potassium ion transport / protein homooligomerization / cytoplasmic side of plasma membrane / cerebral cortex development / lamellipodium / presynaptic membrane / postsynaptic membrane / perikaryon / transmembrane transporter binding / postsynaptic density / cytoskeleton / endosome / neuron projection / protein heterodimerization activity / axon / neuronal cell body / glutamatergic synapse / dendrite / protein-containing complex binding / endoplasmic reticulum membrane / membrane / plasma membrane / cytosol 類似検索 - 分子機能 | ||||||
生物種 | ![]() ![]() | ||||||
手法 | 電子顕微鏡法 / 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 3.3 Å | ||||||
![]() | Matthies, D. / Bae, C. / Fox, T. / Bartesaghi, A. / Subramaniam, S. / Swartz, K.J. | ||||||
![]() | ![]() タイトル: Single-particle cryo-EM structure of a voltage-activated potassium channel in lipid nanodiscs. 著者: Doreen Matthies / Chanhyung Bae / Gilman Es Toombes / Tara Fox / Alberto Bartesaghi / Sriram Subramaniam / Kenton Jon Swartz / ![]() 要旨: Voltage-activated potassium (Kv) channels open to conduct K ions in response to membrane depolarization, and subsequently enter non-conducting states through distinct mechanisms of inactivation. X- ...Voltage-activated potassium (Kv) channels open to conduct K ions in response to membrane depolarization, and subsequently enter non-conducting states through distinct mechanisms of inactivation. X-ray structures of detergent-solubilized Kv channels appear to have captured an open state even though a non-conducting C-type inactivated state would predominate in membranes in the absence of a transmembrane voltage. However, structures for a voltage-activated ion channel in a lipid bilayer environment have not yet been reported. Here we report the structure of the Kv1.2-2.1 paddle chimera channel reconstituted into lipid nanodiscs using single-particle cryo-electron microscopy. At a resolution of ~3 Å for the cytosolic domain and ~4 Å for the transmembrane domain, the structure determined in nanodiscs is similar to the previously determined X-ray structure. Our findings show that large differences in structure between detergent and lipid bilayer environments are unlikely, and enable us to propose possible structural mechanisms for C-type inactivation. | ||||||
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-検証レポート
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集合体
登録構造単位 | ![]()
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要素
#1: タンパク質 | 分子量: 37339.059 Da / 分子数: 4 / 変異: cytosolic domain (UNP residues 37-367) / 由来タイプ: 組換発現 / 由来: (組換発現) ![]() ![]() ![]() #2: タンパク質 | 分子量: 58905.828 Da / 分子数: 4 / 由来タイプ: 組換発現 / 由来: (組換発現) ![]() ![]() ![]() #3: 化合物 | ChemComp-NAP / #4: 水 | ChemComp-HOH / | |
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-実験情報
-実験
実験 | 手法: 電子顕微鏡法 |
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EM実験 | 試料の集合状態: PARTICLE / 3次元再構成法: 単粒子再構成法 |
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試料調製
構成要素 | 名称: Voltage-activated potassium channel Kv1.2-2.1 paddle chimera in lipid nanodiscs タイプ: COMPLEX / Entity ID: #1-#2 / 由来: RECOMBINANT | ||||||||||||||||||||
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分子量 | 値: 0.385 MDa / 実験値: NO | ||||||||||||||||||||
由来(天然) | 生物種: ![]() ![]() | ||||||||||||||||||||
由来(組換発現) | 生物種: ![]() | ||||||||||||||||||||
緩衝液 | pH: 7.5 | ||||||||||||||||||||
緩衝液成分 |
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試料 | 濃度: 0.7 mg/ml / 包埋: NO / シャドウイング: NO / 染色: NO / 凍結: YES / 詳細: Kv1.2-2.1 paddle chimera in lipid nanodiscs | ||||||||||||||||||||
試料支持 | グリッドの材料: GOLD / グリッドのタイプ: Quantifoil, UltrAuFoil, R1.2/1.3 | ||||||||||||||||||||
急速凍結 | 装置: LEICA EM GP / 凍結剤: ETHANE / 湿度: 88 % / 凍結前の試料温度: 277.15 K 詳細: A 3 microliter sample was applied to a plasma-cleaned grid and blotted for 10 seconds. |
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電子顕微鏡撮影
実験機器 | ![]() モデル: Titan Krios / 画像提供: FEI Company |
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顕微鏡 | モデル: FEI TITAN KRIOS |
電子銃 | 電子線源: ![]() |
電子レンズ | モード: BRIGHT FIELD / 倍率(公称値): 29000 X / 最大 デフォーカス(公称値): 2500 nm / 最小 デフォーカス(公称値): 1000 nm / Cs: 2.7 mm / C2レンズ絞り径: 70 µm / アライメント法: COMA FREE |
試料ホルダ | 凍結剤: NITROGEN 試料ホルダーモデル: FEI TITAN KRIOS AUTOGRID HOLDER |
撮影 | 平均露光時間: 15.2 sec. / 電子線照射量: 40 e/Å2 / 検出モード: SUPER-RESOLUTION フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 撮影したグリッド数: 1 / 実像数: 3085 |
画像スキャン | 横: 7420 / 縦: 7676 / 動画フレーム数/画像: 38 / 利用したフレーム数/画像: 2-20 |
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解析
EMソフトウェア |
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CTF補正 | タイプ: PHASE FLIPPING ONLY | ||||||||||||||||||||||||||||||||||||||||
粒子像の選択 | 選択した粒子像数: 281021 | ||||||||||||||||||||||||||||||||||||||||
対称性 | 点対称性: C4 (4回回転対称) | ||||||||||||||||||||||||||||||||||||||||
3次元再構成 | 解像度: 3.3 Å / 解像度の算出法: FSC 0.143 CUT-OFF / 粒子像の数: 47482 / クラス平均像の数: 1 / 対称性のタイプ: POINT | ||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | B value: 89 / プロトコル: FLEXIBLE FIT / 空間: REAL | ||||||||||||||||||||||||||||||||||||||||
原子モデル構築 | PDB-ID: 2R9R Accession code: 2R9R / Source name: PDB / タイプ: experimental model |